Methods and compositions for improving cognitive function

ABSTRACT

This invention relates to compositions, and methods of use thereof, for (i) enhancing executive cognitive function(s) (for example, decision making, planning, working memory, multitasking, judgment, numerical problem-solving, reading comprehension), and/or (ii) increasing blood flow in brain vasculature, comprising administering to a subject in need thereof, certain polyphenols such as flavanols, procyanidins, or pharmaceutically acceptable salts or derivatives thereof.

This application claims the benefit, under 35 USC Section 119, of theU.S. Provisional Appl. No. 60/813,948 filed Jun. 15, 2006, thedisclosure of which is hereby incorporated herein by reference.

FIELD OF THE INVENTION

The invention relates to compositions, and methods of use thereof, for(i) enhancing executive cognitive functions (for example, decisionmaking, planning, working memory, multitasking, judgment, numericalproblem-solving, reading comprehension), and/or (ii) increasing bloodflow in brain vasculature, comprising administering to a subject in needthereof, certain polyphenolic compounds described herein.

BACKGROUND OF THE INVENTION

The flavanols and procyanidins have attracted a great deal of attentionin the fields of medicine and nutrition due to the wide range of theirbiological activities (e.g. U.S. Pat. No. 6,297,273). Applicants havenow discovered that administration of the compounds recited herein leadsto an enhancement of executive cognitive function(s) and/or to anincrease in blood flow in brain vasculature.

Magnetic resonance imaging (MRI) is an imaging technique that is basedon the principles of nuclear magnetic resonance (NMR), a spectroscopictechnique used to obtain microscopic chemical and physical informationabout molecules. MRI is used primarily in medical settings to producehigh quality images of the inside of the human body. MRI systems canalso image flowing blood in virtually any part of the body. This allowsfor studies that show the arterial system in the body, but not thetissue around it. In many cases, the MRI system can do this without acontrast injection, which is required in vascular radiology.

Changes in blood flow and blood oxygenation (collectively known ashemodynamics) in the brain vasculature are known to be closely linkedto/accompany neural activity (i.e., brain activity/function). Therefore,brain activity/function can be studied by mapping changes in brainhemodynamics using imaging techniques. Functional Magnetic ResonanceImaging (fMRI) is one such magnetic resonance (MR)-based technique usedto determine brain function. It measures changes in brain activitythrough the blood oxygenation level dependent contrast (“BOLD”) effect.The BOLD effect is based on changes in distribution of oxygenatedhemoglobin. When brain cells are active they consume oxygen carried byhemoglobin in red blood cells from local capillaries. This oxygenutilization leads to an increase in blood flow to regions of increasedbrain activity which results in local changes in the relativeconcentration of oxyhemoglobin and deoxyhemoglobin. Hence, theoxygenation change measured by the BOLD response arises from a complexinterplay of increases in cerebral blood flow (CBF), cerebral bloodvolume (CBV), and cerebral metabolic rate of oxygen consumption (CMR0₂).Thus, changes in the BOLD signal are well correlated with changes inblood flow, i.e., an increase in the BOLD response is an indication ofincreased blood flow which in turn may correspond to increased cognitivefunction, for example enhanced mental acuity and/or abilities.

Arterial Spin Labeling (ASL) is another non-invasive magnetic resonance(MR)-based imaging technique that provides a physiologically relevantmeasure of CBF in absolute units (i.e., ml of blood/100 grams oftissue/minute). ASL-based imaging is made possible by the same regionalneurovascular coupling that is the basis of BOLD response; however, itprovides a quantitative measure of changes in blood flow itself ratherthan of changes in blood oxygenation.

The use of the above brain imaging techniques (fMRI BOLD response andASL) has allowed for the invention described herein.

Executive cognitive functions play critical roles in the performance ofnumerous complex tasks, for example, decision making, planning, workingmemory, multitasking, judgment, numerical problem-solving, and readingcomprehension. Therefore, there is a need in the art for methods ofenhancing executive cognitive function(s) and/or for increasing bloodflow in the brain.

SUMMARY OF THE INVENTION

This invention relates to compositions, and methods of use thereof, for(i) enhancing executive cognitive function(s), and/or (ii) increasingblood flow in brain vasculature, comprising administering to a subjectin need thereof certain polyphenolic compounds described herein.

In one aspect, the invention relates to a composition, such as apharmaceutical, a food, a food additive, or a dietary supplementcomprising the compounds of the invention. The composition mayoptionally contain an additional cognition-enhancing/improving agent, ormay be administered in combination with an additionalcognition-enhancing/improving agent. Packaged products containing theabove-mentioned compositions and a label and/or instructions for use toenhance/improve mental cognition and/or treat/prevent conditions thatare associated with declining mental cognition are also within the scopeof the invention.

In a further aspect, the invention relates to method of increasing brainactivity (assessed for example by using the fMRI BOLD response) incertain regions of the brain comprising administering the compounds ofthe invention to a subject in need thereof. Non-limiting examples ofsuch activated brain regions include the right-dorsolateral prefrontalcortex, the parietal cortex, and the anterior cingulate.

In another aspect, the invention relates to a method of increasing bloodflow to the brain (measured for example by using ASL) comprisingadministering the compounds of the invention to a subject in needthereof.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1: shows mean reaction time (±SEM) for the letter-digit task.Reaction times were averaged over all subjects.

FIG. 2A: shows task-related activity in the ‘switch’ versus baselinecondition for ‘high flavanol’. Statistical parametric maps thresholdedat P<0.05 (corrected) for height and spatial extent.

FIG. 2B: shows task-related activity for the comparison between ‘switch’and ‘non-switch’ conditions (‘switch’ versus baseline>‘non-switch’versus baseline) for ‘high flavanol’. Statistical parametric mapsthresholded at P<0.05 (corrected) for height and spatial extent.

FIG. 3: shows the time course of the mean cerebral blood flow responses(±SEM) across grey matter (n=4) following ingestion of an acute dose ofhigh flavanol drink and low flavanol drink.

FIG. (4A-4D): shows the effects of a cocoa drink supplemented withflavanols/procyanidins on a “Cognitive Demand Battery” of tests:

FIG. 4A: Threes Correct test;

FIG. 4B: Sevens Error test;

FIG. 4C: Rapid Visual Information Processing Task (RVIP); and

FIG. 4D: Mental fatigue.

DETAILED DESCRIPTION

All patents, patent applications and references cited in thisapplication are hereby incorporated herein by reference. In case of anyinconsistency, the present disclosure governs.

The present invention relates to compositions, products, and methods for(i) enhancing executive cognitive function(s), and/or (ii) increasingblood flow in brain vasculature, each comprising administering, to asubject in need thereof, certain polyphenolic compounds describedherein. The compounds for use in the present invention include certainflavanols (flavan-3-ols), procyanidins, or pharmaceutically acceptablesalts or derivatives thereof. Such compounds, when of natural origin,may be included in the composition in the form of a cocoa component, forexample cocoa nibs or fragments thereof, chocolate liquor, cocoa solids(e.g. partially or fully-defatted), cocoa extract or fraction thereof.

As used herein, the term “flavanol” or “flavan-3-ol” refers to a monomerof the following formula:

The term “procyanidin” refers to an oligomer of the monomer shown above.

The term “cocoa component” refers to a component derived from cocoabean, e.g. cocoa nibs and fragments thereof, chocolate liquor, cocoasolids (e.g. partially and fully-defatted cake or powder), flavanoland/or procyanidin-containing cocoa extract or fraction thereof.

In certain embodiments, the present invention relates to a flavanol(e.g. (−)-epicatechin and (+)-catechin), and a composition comprising aneffective amount of the flavanol (e.g. (−)-epicatechin and(+)-catechin), or a pharmaceutically acceptable salt or derivativethereof (including oxidation products, methylated derivatives, andglucuronidated derivatives). The derivatives may be prepared asdescribed below.

In other embodiments, the present invention relates to a compound, and acomposition comprising an effective amount of the compound, having thefollowing formula An, or a pharmaceutically acceptable salt orderivative thereof (including oxidation products, methylatedderivatives, and glucuronidated derivatives):

wherein

-   -   n is an integer from 2 to 18;    -   R and X each have either a or stereochemistry;    -   R is OH or 0-sugar or 0-gallate;    -   the substituents of C-4, C-6 and C-8 are X, Z and Y,        respectively, and bonding of monomeric units occurs at C-4, C-6        or C-8;    -   when any C-4, C-6 or C-8 is not bonded to another monomeric        unit, X, Y and Z independently are hydrogen or a sugar; and    -   the sugar is optionally substituted with a phenolic moiety at        any position, for instance, via an ester bond.

Monomeric units in the formula An may be bonded via 4 - - - +6a; 4 - - -+6; 4 - - - +8a; and/or 4 - - - +8 linkages. The sugar is preferably amonosaccharide or a disaccharide. The sugar may be selected from thegroup consisting of glucose, galactose, rhamnose, xylose, and arabinose.The phenolic moiety may be selected from the group consisting ofcaffeic, cinnamic, coumaric, ferulic, gallic, hydroxybenzoic and sinapicacids. Derivatives may include esters such as the gallate esters;compounds derivatized with a saccharide moiety such as mono- ordisaccharide moiety (e.g. -D-glucose), glucuronidated and methylatedderivatives, and oxidation products. In some embodiments, esterderivatives are other than esters with gallic acid. Oxidation productsmay be prepared as disclosed in U.S. Pat. No. 5,554,645, the relevantportions of which are incorporated herein by reference. Esters, forexample esters with gallic acid, may be prepared using knownesterification reactions, and for example as described in U.S. Pat. No.6,420,572, the disclosure of which is hereby incorporated herein byreference. Methylated derivatives, such as 3′0-methyl-, 4′0-methyl-, and3′O,4′0-dimethyl-derivatives may be prepared, for example, as describedin Cren-Olive et al., 2002, J. Chern. Soc. Perkin Trans. 1, 821-830, andDonovan et al., Journal of Chromatography B, 726 (1999) 277-283, thedisclosures of which are hereby incorporated herein by reference.Glucuronidated products may be prepared as described in Yu et al, “Anovel and effective procedure for the preparation of glucuronides,”Organic Letters, 2 (16) (2000) 2539-41, and as in Spencer et al,“Contrasting influences of glucuronidation and 0-methylation ofepicatechin on hydrogen peroxide-induced cell death in neurons andfibroblasts,” Free Radical Biology and Medicine 31 (9) (2001) 1139-46,hereby incorporated herein by reference. It should be noted that thisdisclosure applies to all formulas recited herein.

In another embodiment, the invention relates to a compound, and thecomposition comprising an effective amount the compound having theformula An, or a pharmaceutically acceptable salt or derivative thereof(including oxidation products, methylated derivatives, andglucuronidated derivatives):

wherein

-   -   n is an integer from 2 to 18;    -   R and X each have either α or stereochemistry;    -   R is OH;    -   the substituents of C-4, C-6 and C-8 are X, Z and Y,        respectively, and bonding of monomeric units occurs at C-4, C-6        and C-8; and    -   when any C-4, C-6 or C-8 is not bonded to another monomeric        unit, X, Y and Z are hydrogen.

Examples of the compounds useful for the products and in the methods ofthe invention include the compounds of the formula An described hereinwherein the integer n is 3 to 18; 2 to 12; 3 to 12; 2 to 5; 4 to 12; 5to 12; 4 to 10; or 5 to 10. In some embodiments, the integer n is 2 to4, for example 2 or 3. This disclosure applies to any compound offormula An herein.

Methods of Use

The invention relates to methods of (i) enhancing executive cognitivefunction(s) and/or (ii) increasing blood flow in brain vasculature.

As used herein, “executive cognitive function” is defined as a higherorder cognitive capacity that plays a role in managing (like an“executive”) other cognitive functions such as attention, language andmemory. Examples of executive cognitive functions includedecision-making, multi-tasking, working memory, performance of complexnumerical calculations, reading comprehension.

As used herein, “enhancement of cognitive function” is defined as ameasurable improvement of at least one of the executive cognitivefunctions. A person of skill in the art will select the known methods ofmeasuring the improvement of cognitive functions, for example, methodsdescribed in the Examples.

As used herein, “decline in cognitive abilities” is defined as adeterioration of cognitive abilities in a healthy subject, for examplean elderly/aged subject, i.e., as used herein “decline in cognitiveabilities” is not referring to a neurodegenerative condition.

As used herein, a “healthy” subject is one who is not suffering from/hasnot been diagnosed with a neurodegenerative disease.

As used herein, “increasing blood flow” refers to an increase in theamount of blood delivered to the tissue which may be expressed in termsof milliliters (ml) of blood per 100 ml of tissue per minute. Increasingblood flow in the brain refers to an increase in the volume of bloodentering a unit volume of brain per unit of time.

In certain embodiments, the present invention provides (i) a method ofenhancing an executive cognitive function and/or (ii) a method ofincreasing blood flow to brain vasculature, each comprisingadministering to a mammal (e.g. human) or a veterinary animal in needthereof an effective amount of a flavanol such as epicatechin orcatechin (e.g. (−)-epicatechin or (+)-catechin), or a pharmaceuticallyacceptable salt or derivative thereof (including oxidation products,methylated derivatives, and glucuronidated derivatives).

The term “veterinary animal” refers to any animal cared for, or attendedto by, a veterinarian, and includes companion (pet) animals andlivestock animals, for example a cat, a dog and a horse (e.g. a racehorse).

In certain embodiments, the invention provides (i) a method of enhancingan executive cognitive function and/or (ii) a method of increasing bloodflow to brain vasculature, each comprising administering to a mammal(e.g. human) or a veterinary animal in need thereof, a compositioncomprising an effective amount of a compound having the followingformula An, or a pharmaceutically acceptable salt or derivative thereof(including oxidation products, methylated derivatives, andglucuronidated derivatives):

wherein

-   -   n is an integer from 2 to 18;    -   R and X each have either α or stereochemistry;    -   R is OH or 0-sugar or 0-gallate;    -   the substituents of C-4, C-6 and C-8 are X, Z and Y,        respectively, and bonding of monomeric units occurs at C-4, C-6        or C-8;    -   when any C-4, C-6 or C-8 is not bonded to another monomeric        unit, X, Y and Z independently are hydrogen or a sugar; and    -   the sugar is optionally substituted with a phenolic moiety at        any position, for instance, via an ester bond.

For example, the above method may involve use of a compound An, or apharmaceutically acceptable salt or derivative thereof (includingoxidation products, methylated derivatives and glucuronidatedderivatives), wherein R is OH, and when any C-4, C-6 or C-8 is notbonded to another monomeric unit, X, Y and Z are hydrogen. Examples ofsuitable sugars are as described above. Examples of phenolic moietiesare as described above. Examples of derivatives are as described above.

In certain embodiments, the invention provides (i) a method of enhancingan executive cognitive function and/or (ii) a method of increasing bloodflow in brain vasculature, each comprising administering to a mammal(e.g. human) or a veterinary animal in need thereof, a compositioncomprising an effective amount of a compound having the formula An, or apharmaceutically acceptable salt or derivative thereof (includingoxidation products, methylated derivatives, and glucuronidatedderivatives):

wherein

-   -   n is an integer from 2 to 18;    -   R and X each have either α or ρ stereochemistry;    -   R is OH;    -   the substituents of C-4, C-6 and C-8 are X, Z and Y,        respectively, and bonding of monomeric units occurs at C-4, C-6        and C-8; and    -   when any C-4, C-6 or C-8 is not bonded to another monomeric        unit, X, Y and Z are hydrogen.

Examples of the compounds useful for the products and in the methods ofthe invention include the compounds described herein wherein the integern is 3 to 18; 2 to 12; 3 to 12; 2 to 5; 4 to 12; 5 to 12; 4 to 10; or 5to 10. In some embodiments, the integer n is 2 to 4, for example 2 or 3.This disclosure applies to any compound of formula An herein.

While any human or a veterinary animal can benefit from the methodsdescribed herein, it will be understood that a “subject in need thereof”is a subject in imminent need of enhancement of cognitive function or asubject having a profession (or performing tasks) that require(s)constant utilization of executive cognitive function(s). Examples ofsubjects in need of enhancement of cognitive function(s) and/orincreased brain blood flow will be apparent to those of skill in theart, for example subjects who will imminently participate in acompetitive event (e.g. those appearing in an examination, competing insporting/athletic events, attending a job interview); those addressingan audience/gathering (e.g. educators, politicians, live television-newsanchors, reporters); entertainers (e.g. movie actors and live-showperformers); and those engaged in tasks involving intense periods ofmental processing. Also included are subjects who perform tasksrequiring complex decision making and multitasking, for examplehigh-profile business executives; transportation workers (e.g. busdrivers, railway engine drivers, airline pilots, shipcaptains/commanders); construction workers (e.g. high-risk constructionssuch as bridges, high-rise buildings, roadways and railways onmountains); defense personnel (e.g. warship commanders, warrior aircraftpilots, pilots of naval vessels). Further, an elderly/aged individual(for example >65, e.g. >75) who is experiencing cognitive decline buthas not been clinically diagnosed with a neurodegenerative disease willalso benefit from the compositions and methods described herein.

The present compounds may be administered orally in the form of a cocoacomponent, for example cocoa nibs or fragments thereof, chocolateliquor, cocoa sol ds (e.g. partially and fully-defatted cocoa solid,e.g. of cocoa solids is cocoa poWder), cocoa extract or fractionthereof, or may be added independently of cocoa components. The cocoacomponent may be prepared such that the content of cocoa polyphenols(CP) is preserved for example by altering traditional processing steps,as described, for example, in U.S. Pat. Nos. 6,194,020 and 6,599,553.

In some embodiments, the present compounds may be administered incombination with other cognition-enhancing agents and/or to enhanceresponsiveness to other cognition-enhancing agents. Examples ofcognition-enhancing agents include: metabolic substrates (e.g., glucose,ketones, supplemental oxygen), alkaloids (e.g., theobromine, caffeine)vitamins, amino acids, minerals, micronutrients, botanical extracts ortheir derivatives, herbs or herbal supplements (e.g., ginkgo, ginseng).

Thus, the following uses are within the scope of the invention. Use of aflavanol, or a pharmaceutically acceptable salt or derivative thereof(including oxidation products, methylated derivatives, andglucuronidated derivatives) as defined above, in the manufacture of amedicament, food, nutraceutical or dietary supplement for enhancing anexecutive cognitive function and/or for increasing blood flow in brainvasculature. Use of a compound of formula An, or a pharmaceuticallyacceptable salt or derivative thereof (including oxidation products,methylated derivatives and glucuronidated derivatives), as definedherein, in the manufacture of a medicament, food, nutraceutical ordietary supplement for use in enhancing an executive cognitive functionand/or for increasing blood flow in brain vasculature.

The effective amount may be determined by a person of skill in the artusing the guidance provided herein and general knowledge in the art. Forexample, the effective amount may be such as to achieve aphysiologically relevant concentration in the body of a mammal. Such aphysiologically relevant concentration may be at least 20 nanomolar(nM), preferably at least about 100 nM, and more preferably at leastabout 500 nM. In one embodiment, at least about one micromole in theblood of the mammal, such as a human, is achieved. The compounds definedherein, may be administered at from about 35 mg/day, 40 mg/day or 50mg/day (e.g. to about 1000 mg/day), or from about 75 mg/day (e.g. toabout 1000 mg/day), or from about 100-150 mg/day (e.g. to about 900mg/day), or from about 300 mg/day (e.g. to about 500 mg/day). However,amounts higher than exemplified above may be used since the upper end ofthe amount range is not a limiting factor. The amounts may be measuredas described in Adamson, G. E. et al., J. Ag. Food Chern.; 1999; 47 (10)4184-4188.

The administration may be continued as a regimen, i.e., for an effectiveperiod of time, e.g., daily, monthly, bimonthly, biannually, annually,or in some other regimen, as determined by the skilled medicalpractitioner for such time as is necessary. The administration may becontinued for at least a period of time required for enhancement ofexecutive cognitive function(s) and/or for increasing blood flow inbrain vasculature. The composition may be administered daily, preferablytwo or three times a day, for example, morning and evening to maintainthe levels of the effective compounds in the body of the mammal. Toobtain the most beneficial results, the composition may be administeredfor at least 7 days, or at least 14 days, or at least 30 days, or atleast 45 days, or at least 60 days, or at least 90 days. These regimensmay be repeated periodically as needed.

Compositions and Formulations

The compounds of the invention may be administered as a food (includingpet food), a food additive, or a dietary supplement, or apharmaceutical.

As used herein, “food” is a material containing protein, carbohydrateand/or fat, which is used in the body of an organism to sustain growth,repair and vital processes and to furnish energy. Foods may also containsupplementary substances, for example, minerals, vitamins andcondiments. See Merriam-Webster's Collegiate Dictionary, 10th Edition,1993. The term food includes a beverage adapted for human or animalconsumption. As used herein a “food additive” is as defined by the FDAin 21 C.F.R. 170.3(e)(1) and includes direct and indirect additives. Asused herein, a “dietary supplement” is a product (other than tobacco)that is intended to supplement the diet that bears or contains one ormore of the following dietary ingredients: a vitamin, a mineral, an herbor other botanical, an amino acid, a dietary substance for use by man tosupplement the diet by increasing the total daily intake, or aconcentrate, metabolite, constituent, extract or combination of theseingredients. As used herein, a “pharmaceutical” is a medicinal drug. SeeMerriam-Webster's Collegiate Dictionary, 10th Edition, 1993. Apharmaceutical may also be referred to as a medicament. The abovecompositions may be prepared as is known in the art.

The compositions may contain a carrier, a diluent, or an excipient.Depending on the intended use, the carrier, diluent, or excipient may bechosen to be suitable for human or veterinary use, food, additive,dietary supplement or pharmaceutical use. The composition may optionallycontain an additional cognition-enhancing/improving agent. Alsodepending on use, a person of skill in the art may select the degree ofpurity of the compound of the invention. For example, when used toprepare pharmaceutical dosage forms, the compound should be as pure ascommercially possible, while when preparing food, additive, orsupplement, less pure or mixtures of compounds (e.g. plant extracts) maybe used.

The compound of the invention may be “isolated and purified,” i.e., itmay be separated from compounds with which it naturally occurs (e.g.when the compound is of natural origin), or it may be syntheticallyprepared, in either case such that the level of contaminating compoundsand/or impurities does not significantly contribute to, or detract from,the effectiveness of the compound. For example, an “isolated andpurified B2 dimer” is separated from BS dimer, with which it may occurin nature (e.g. in cocoa bean), to the extent achievable by theavailable commercially viable purification and separation techniques.Such compounds are particularly suitable for pharmaceuticalapplications.

The compound may also be less pure, i.e., “substantially pure,” i.e., itmay possess the highest degree of homogeneity achievable by availablepurification, separation and/or synthesis technology but need not beseparated from the like compounds. As used herein, “the like compounds”are the compounds having the same degree of polymerization. For example,a “substantially pure dimer” refers to a mixture of dimers (e.g. B2 andB5, as it would occur in a cocoa extract fraction). While less suitablefor pharmaceutical applications, such “substantially pure” compounds maybe utilized for food, food additive and dietary supplement applications.

In some embodiments, the compound of the invention is at least 80% pure,at least 85% pure, at least 90% pure, at least 95% pure, at least 98%pure, or at least 99% pure. Such compounds are particularly suitable forpharmaceutical applications.

Pharmaceuticals containing the inventive compounds, optionally incombination with another cognition-enhancing/improving agent may beadministered orally. As used herein, “oral administration” includesadministration by the mouth and includes sublingual and buccaladministrations. A person of skill in the art will be able to determinea suitable mode of administration to maximize the delivery of thecompounds of the invention. Thus, dosage forms adapted for each type ofadministration by mouth are within the scope of the invention andinclude solid, liquid and semi-solid dosage forms, such as tablets,capsules, gelatin capsules (gelcaps), bulk or unit dose powders orgranules, emulsions, suspensions, pastes, or jellies. Sustained-releasedosage forms are also within the scope of the invention. Suitablepharmaceutically acceptable carriers, diluents, or excipients aregenerally known in the art and can be determined readily by a personskilled in the art. The tablet, for example, may comprise an effectiveamount of the compound of the invention and optionally a carrier, suchas sorbitol, lactose, cellulose, or dicalcium phosphate.

The foods comprising the compounds described herein and optionallyanother cognition-enhancing/improving agent may be adapted for human orveterinary use, and include pet foods. The food may be other than aconfectionery, for example, a beverage (e.g. cocoa flavored beverage). Aconfectionery such as a standard of identity (SOI) and non-SOIchocolate, such as milk, sweet and semi-sweet chocolate including darkchocolate, low fat chocolate and a candy which may be a chocolatecovered candy are also within the scope of the invention. Other examplesinclude a baked product (e.g. brownie, baked snack, cookie, biscuit) acondiment, a granola bar, a toffee chew, a meal replacement bar, aspread, a syrup, a powder beverage mix, a cocoa or a chocolate flavoredbeverage, a pudding, a rice cake, a rice mix, a savory sauce and thelike. If desired, the foods may be chocolate or cocoa flavored. Foodproducts may be chocolates and candy bars, such as granola bars,containing nuts, for example, peanuts, walnuts, almonds, and hazelnuts.The food is designed to deliver an effective amount of the compoundsdescribed herein.

The compounds for use in the present invention may be of natural origin,for example, derived from a cocoa bean or another natural source knownto a person of skill in the art, or prepared synthetically. A person ofskill in the art may select natural or synthetic polyphenols based onthe use and/or availability or cost.

The compounds may be included in the composition in the form of a cocoacomponent. For example, the compound(s) may be included in thecomposition in the form of a cocoa ingredient, for example, chocolateliquor included in chocolate, or may be added independently of cocoaingredients, for example, as an extract, extract fraction, isolated andpurified individual compound, pooled extract fractions or asynthetically prepared compound. The extraction and purification may beconducted as described in U.S. Pat Nos. 5,554,645 and 6,670,390 toRomanczyk et al., and U.S. Pat. No. 6,627,232 to Hammerstone et al.,each of which is hereby incorporated herein by reference.

Cocoa flavanols and/or procyanidins may be provided in the compositionof the invention by cocoa ingredients containing these compounds or byincluding chocolate, which may be milk, sweet and semi-sweet, and ispreferably dark chocolate, and low fat chocolate. The cocoa ingredientsmay be prepared using traditional cocoa processing procedures but ispreferably prepared using the method described in U.S. Pat. No.6,015,913 to Kealey et al. Alternatively, to enhance the level of cocoapolyphenols, chocolate liquor and cocoa solids prepared from cocoa beanshaving a fermentation factor of 275 or less may be used. Theseingredients have cocoa polyphenol content that is higher than can beobtained using traditional cocoa processing methods (e.g. with roastingand fully fermented beans). The chocolate may be prepared usingconventional techniques from the ingredients described above or using animproved process for preserving cocoa polyphenols during chocolatemanufacturing as described in the International Appl. No. PCT/US99/05414published as W099/45788 and in its U.S. counterpart, U.S. Pat. No.6,194,020, the relevant portions of which are hereby incorporated hereinby reference. A chocolate prepared by at least one of the followingnon-traditional processes is referred to herein as a “chocolate having aconserved amount of cocoa polyphenols”: (i) preparing cocoa ingredientsfrom underfermented or unfermented cocoa beans; (ii) preserving cocoapolyphenol during cocoa ingredient manufacturing process; and (iii)preserving cocoa polyphenol during chocolate manufacturing process. Suchnon-traditional processes may be used to prepare other cocoacomponent-containing products (foods e.g. beverages, dietarysupplements) designed to contain enhanced levels of flavanols and/orprocyanidins.

Synthetic procyanidins may also be used and are prepared by methodsknown in the art and as described, for example, as in U.S. Pat. Nos.6,420,572; 6,156,912; and 6,864,377, the relevant portions of each ofwhich are hereby incorporated herein by reference.

A daily effective amount of the compound of the invention may beprovided in a single serving in case of a food or a single dosage incase of a pharmaceutical or a dietary supplement. For example, aconfectionery (e.g. chocolate) may contain at least about 100 mg/serving(e.g. 150-200, 200-400 mg/serving).

The dietary supplement containing the compounds of the invention, andoptionally another cognition-enhancing/improving agent, may be preparedusing methods known in the art and may comprise, for example, nutrientsuch as dicalcium phosphate, magnesium stearate, calcium nitrate,vitamins, and minerals.

Further within the scope of the invention is an article of manufacturesuch as a packaged product comprising the composition of the invention(e.g. a food, a dietary supplement, a pharmaceutical) and a labelindicating the presence of, or an enhanced content of the inventivecompounds, or directing use of the composition to enhance executivecognitive function(s) and/or to increase blood flow in brainvasculature. The packaged product may contain the composition and theinstructions for use to enhance executive cognitive functions and/or toincrease blood flow in brain vasculature. The label and/or instructionsfor use may refer to any of the methods of use described in thisapplication.

The invention also relates to a method of manufacturing an article ofmanufacture comprising any of the compositions described herein,packaging the composition to obtain an article of manufacture andinstructing, directing or promoting the use of the composition/articleof manufacture for any of the uses described herein. Such instructing,directing or promoting includes advertising.

The invention is further described in the following non-limitingexamples.

EXAMPLES Example 1: Effect of Cocoa Flavanols on the fMRI Response to aCognitive Task in Healthy Young People Methods Subjects

Sixteen young female subjects between the ages of 18 and 30 yrparticipated in the study under the following exclusion criteria: nohistory of migraines, stroke, hypertension, diabetes, or anyneurological or vascular disease and no use of tobacco products. Allsubjects had normal vision and normal color vision, were not dyslexic,were right handed and their first language used the ‘Roman’ alphabet.Subjects average daily caffeine intake was estimated from theirresponses to a dietary questionnaire and they were all classified as lowcaffeine users (<120 mg/day). All subjects were instructed to refrainfrom alcohol and caffeine, or from using any medication for 12 hrsbefore each visit for the fMRI measurements. The local Medical SchoolResearch Ethics Committee approved this study and all subjects gaveinformed written consent before taking part in the study.

Each subject underwent two fMRI sessions that were repeated at least 14days apart. Subjects were randomized to receive a high flavanol cocoadrink (150 mg flavanollprocyanidins per drink for 5 days prior to onefMRI session and a low flavanol cocoa drink (13 mg flavanol/procyanidinper drink) for 5 days prior to the other session in a double blindcounterbalanced manner. These will be referred to hereafter as ‘highflavanol’ and ‘low flavanol’. Subjects consumed one drink per day at aset time for the 5 days prior to each scan session, with the final drinkbeing consumed approximately 1.5 hours before the fMRI scan.

Study Design

Subjects were pre-trained to perform two tasks, a number task ofodd-even judgment and a letter task requiring consonant-vowel judgment.In the letter task subjects learnt to respond to single letter stimulidepending on whether the letter displayed was a consonant (G, K, M, R)or a vowel (A, E, O, U). Subjects were trained to press a left button inresponse to a vowel and right button for a consonant. In the numbertask, subjects were trained to respond to digits that were either odd(3, 5, 7, 9) or even (2, 4, 6, 8), using left and right button responsesrespectively.

Once subjects were familiarized with the rules of the letter and numbertasks, they were trained on the letter-digit pairs task to be performedin the fMRI study. The letter-digit pairs task consisted of a letter anda digit displayed simultaneously on a computer screen (for the fMRIscanning a projector and screen were used). The letter-digit pairs wereeither red or blue. When letter-digit pairs were presented in red,subjects were instructed to attend to the letter and respond by pressingthe appropriate button as trained (i.e., applying the rule forcategorizing as vowel or consonant). If the letter-digit pair was bluethey responded to the digit (odd-even judgment) in a similar manner.

In this study the definition of the ‘switch’ task is the changingbetween the two sets of rules, one for the letters (consonant-voweljudgment) and one for digits (odd-even judgment). To create a paradigmcomprising of ‘switch’ and ‘non-switch’ conditions, the letter-digitpairs were grouped into blocks. A block of five letter-digit pairs, allof the same color is a ‘non-switch’ block. A block of five letter-digitpairs alternating between red and blue stimuli (and so reconfiguringtask judgment) is a ‘switch’ block. The gap between each letter-digitpair within the block was 3 seconds, giving a total block length of 15seconds; this was then followed by a 12 second fixation cross (baselinecondition). The blocks were presented alternately (i.e., ‘switch’ block,‘non-switch’ block, ‘switch’ block, ‘non-switch’ block etc.). Thepresentation of letter-digit pairs within a block helps to increase theswitch cost and so also increase the magnitude of the fMRI BOLDresponse, whilst the 12 second interval between the blocks allows theBOLD response to return to baseline.

Prior to the fMRI study the subjects were trained to a competent level(error rates below 5%) in the task by performing five blocks of ‘switch’and ‘non-switch’ trials. During the fMRI study twenty blocks of ‘switch’trials and 20 blocks of ‘non-switch’ trials were performed in the study,resulting in a total study duration of 18 minutes.

jMRI Scanning

A 3.0 T purpose-built scanner was used with TEM head coil and inserthead gradient coil. T2*-weighted coronal echo-planar images (EPI) with a128×64 matrix size, 3 mm in-plane resolution and 9 mm slice thicknesswere acquired using MBEST acquisition sequence with 30 ms echo time (TE)and 1.9 kHz gradient switching frequency. Sixteen contiguous coronalslices were acquired every 3 seconds (TR=3 s). Throughout the studysubjects reaction time and error rate data was recorded. Further thesubjects' heart rate was monitored. Following the fMRI study a 64 sliceEPI set was acquired to aid anatomical localization.

jMRI Data Processing

The fMRI data was processed using SPM99 (Friston K J et al., Neuroimage1995; 2:166-172) (Statistical Parametric Mapping, Wellcome Department ofImaging Neuroscience, UK). The raw data from the scanner was motioncorrected to realign all functional slices to the first volume of thedata set, spatially normalized to the standard EPI template. Eightmillimeter FWHM (full width half maximum) spatial smoothing and 128 shigh pass filter cut-off were applied.

A general linear model design matrix was created within SPM99 thatmodelled the paradigm. ‘Switch’ and ‘non-switch’ blocks were modelled as12 s box functions. The paradigm time course was then convolved with thecanonical haemodynamic response function and its temporal derivative.Statistical parametric maps (SPM's) of the ‘switch’ condition and‘non-switch’ condition vs baseline were formed. In addition, directcomparison between the two activation conditions (i.e., ‘switch’ vs.baseline and ‘non-switch’ vs. baseline) was also performed at acorrected significance level of P<0.05, with the respective activationvs. baseline comparisons (‘switch’ vs baseline and ‘non-switch’ vsbaseline) being used as a mask.

Cerebral Blood Flow (CBF) Measurements and Analysis

In this initial study on four subjects (24 to 31 yr) we assessed thetime course of the effect of flavanols on brain blood flow. A cerebralblood flow (CBF) map was acquired using an EPISTAR (Echo-Planar MRImaging and Signal Targeting with Alternating Radiofrequency) arterialspin labelling sequence (ASL) on five multi-slice axial 7 mm slices.Diffusion weighting (b=5 mm²/s) was applied for suppression ofintra-arterial spins. A hyperbolic secant pulse was used for thelabelling, and the tag and control slabs were 9 cm in width with aninversion time (TI) of 1400 ms. The EPISTAR sequence was implementedwith repetition time (TR) of 3 seconds between tag and control images,and a total of 60 tag and control pairs were acquired. Each subjectunderwent CBF imaging prior to and at 2, 4, and 6 hours after ingestionof a high flavanol cocoa drink (450 mg flavanol) or a low flavanol cocoadrink, on two separate occasions. In this study, in contrast to the fMRIstudy, a single dose of the drinks was consumed on only one occasion.Each CBF measurement was followed by the acquisition of a T₁ map forsegmentation of brain tissue types and grey matter territories. (CBV iscommonly used to refer to blood volume).

Cerebral blood flow data were first segmented into grey and white matterregions using masks generated from the T₁ map. Grey matter CBF maps werethen further segmented into territories fed by major vessels (Yen Y-F etal., Magn Reson Med 2001: 47:921-928; hereby incorporated herein byreference). Mean cerebral blood flow values were then calculated forwhite and grey matter regions. Whole grey matter perfusion values arepresented here.

Results Behavioural Results for Task Switching Paradigm

Robust switch costs in response times were observed. Each of thesubjects was numerically slower for the ‘switch’ condition than‘non-switch’ condition, both for the letter and number tasks, theconstant switching from one task rule to the other proving difficult.The mean reaction times across the group for the ‘switch’ and‘non-switch’ blocks are shown in FIG. 1. The significance of the switchcost was p=5×10-⁶ for ‘low flavanol’ and p=1×10-⁶ for ‘high flavanol’.There was no significant difference in the switch cost between the twodrinks (p=0.30). The average switch cost was 224±25 milliseconds (ms).

The possibility that during the 40 blocks of ‘switch’ and ‘non-switch’trials of the fMRI study the subjects would either make more or lesserrors as they either became fatigued or improved due to learningeffects was also investigated. Analysis of the reaction time responsesrevealed that no significant fatigue/learning effects occurred over thecourse of the fMRI sessions (p=0.74). Also the drink order wasrandomized and a comparison of the reaction time responses between firstand second scanning sessions made, this again revealed no significantdifferences (p=0.73).

Heart Rate Results for Task Switching Paradigm

The mean heart rate for the ‘switch’ and ‘non-switch’ conditions for thelow and high flavanol cocoa drinks was measured as shown in Table 1.Paired t-tests were performed to determine any differences in heart ratebetween the ‘switch’ and ‘non-switch’ conditions; for both drinks therewas a significant increase in heart rate for the ‘switch’ conditioncompared to ‘non-switch’ condition (‘low flavanol’:‘switch’>‘non-switch’ p=0.01; ‘high flavanol’: switch’>‘non-switch’p=0.0009). No significant difference in heart rate was found between thelow and high flavanol drinks.

TABLE 1 The mean heart rate(±SEM) (beats per minute) in response to‘switch’ and ‘non-switch’ conditions for low and high flavanol drinks.Low flavanol High flavanol Switch 66.8 +/− 2.6 67.6 +/− 2.6 Non-switch63.0 +/− 2.5 64.2 +/− 3.0

jMRI Results for Task Switching Paradigm

FIG. 2a shows the group statistical parametric map for the switch taskvs baseline condition at a corrected probability of P<0.05. The ‘switch’and ‘non-switch’ versus baseline conditions revealed activation in themedial and lateral prefrontal cortex (including the dorsolateralprefrontal cortex (DLPFC)), parietal cortex, anterior cingulate cortex(ACC), and cerebellum. ‘Non-switch’ is not represented in FIG. 2a ,however, ‘non-switch’ vs baseline condition shows similar areas to the‘switch’ vs baseline condition.

The brain areas outlined above have previously been shown to beassociated with task switching (Lewis P A et al., Curr OpinNeurobio/2003; 13:250-255; Sohn M H et al., Proc Nat/Acad Sci USA 2000;97:13448-53; Swainson R et al., J Cogn Neurosci 2003; 15:785-99).Further, a number of cognitive neuroimaging studies aside fromtask-switching have found similar patterns of activation. These includeworking memory (Cabeza et al., Neuroimage 2002: 16:317-330; Postle B etal., Proc Nat/Acad Sci USA 1999; 96:12959-12964), memory retrieval(Thompson-Schill S L et al., Proc Nat/Acad Sci USA 1997: 94:14792-14797;Wagner A D et al., Neuroimage 2000; 14:1337-1347), and arithmeticproblem solving tasks (Anderson J R et al., Psychonom Bull Rev 2003;10:241-261; Dehaenae S et al., Science 1999; 284:970-974). All suchtasks demonstrate strong prefrontal-parietal interconnections (PetridesMetal., J Comp Neuro/1984; 228:105-116; Schwartz M L et al., J CompNeuro/1984; 226:403-420), suggesting that these two areas may servecomplementary roles in the high-level cognition.

FIG. 2b shows the group statistical map of areas of activation whichshow significantly increased BOLD response during the ‘switch’ taskrelative to the ‘non-switch’ task. From this comparison, it can be seenthat those brain areas activated preferentially to the ‘switch’condition are largely localized in the right hemisphere, in thedorsolateral prefrontal and parietal cortices, as well as the anteriorcingulate cortex and cerebellum.

FIG. 2a shows areas for the ‘switch’ condition relative to a restingbaseline. These are all those parts of the brain which are associatedwith performing the switch task, so are not exclusive to task switchingsince they can include areas associated with different aspects of thetask such as the motor response to the button press.

Referring to FIGS. 2a-b , FIG. 2b shows areas which are greater for the‘switch’ condition than the ‘non-switch’ condition. Therefore, this canbe thought of as (‘switch’ condition relative to a restingbaseline)−(‘non-switch’ condition relative to a resting baseline). Thisis the important condition as the aspects of the task which areunimportant, such as the motor response to the button press, are in boththe ‘switch’ and ‘non-switch’ conditions and so by subtracting‘non-switch’ from ‘switch’ those brain areas associated with the motorbutton press response are removed. In the ‘switch’ minus ‘non-switch’condition only those areas which are purely involved in task switchingare seen.

In task-switching it is thought that the anterior cingulate cortex (ACC)detects conflict in a task-setting (Gehring W J and Knight R T, NatNeurosci 2000; 3:516-20), the right frontal cortex plays a role relatedto the inhibition of irrelevant (preceding) responses (Aron et a/.,Trends Cogn Sci 2004:8:170-7), and the active maintenance of informationthat is newly loaded into working memory (Goldman-Rakic P S, Annu RevNeurosci 1988; 11:137-156). The right posterior parietal cortex haswidely been shown to be responsible for spatial or visual attention(Rushworth M F S eta/., J Neurosci 2001; 21: 5262-5271), whilst thecerebellum is thought to be primarily activated with timing irregularityin the switch task, consistent with its role as an internal timingsystem (Ivry R B, Curr Opin Neurobio/1996; 6:851-7).

Statistical comparison of the BOLD signal change between the ‘lowflavanol’ and ‘high flavanol’ conditions revealed that the ‘highflavanol’ generated a significantly greater BOLD signal change for boththe activation (‘switch’ and ‘non-switch’) versus baseline conditions aswell as for the comparison of ‘switch’ vs baseline with ‘non-switch’ vsbaseline condition. Table 2 shows the average percentage signal changefor the ‘switch’ BOLD response relative to baseline, for selectedregions of interest, following ingestion of ‘low’ and ‘high’ flavanoldrinks.

TABLE 2 The average percentage signal change(±SEM) of the BOLD responserelative to baseline for the ‘switch’ condition following ingestion of arepeated dose of low and high flavanol drinks. The high flavanol drinkrevealed a marked increase in the BOLD response. Low flavanol Highflavanol Dorsolateral prefrontal cortex 2.3 ± 0.2 3.0 ± 0.2 (DLPFC)Parietal cortex 2.1 ± 0.3 2.5 ± 0.2 Anterior Cingulate Cortex 1.7 ± 0.12.1 ± 0.3 (ACC)

Cerebral Blood Flow (CBF) Time Course Results

FIG. 3 shows the time course of the mean cerebral blood flow responseacross grey matter following ingestion of the low and high flavanoldrinks. It can be seen that there was an increase in cerebral blood flowin response to the high flavanol drink, with a peak in the cerebralblood flow response occurring at approximately two hours post ingestion,and CBF returning to baseline after approximately six hours. It shouldbe noted that in this study an acute dose of flavanols was given, incontrast to the repeated dose given in the tMRI study.

ASL studies have previously been performed to measure the effects ofhypercapnia, which induces cerebral vasodilation and CBF increases. Suchstudies have shown a CBF increase from 30% up to 87% in response to anexpiration breath hold, which leads to an instantaneous increase inPaCOz (Li T Q eta/., Neuroimage 1999; 10:562-9; Li T Q et al.,Neuroimage 1999; 9:243-9; Kastrup A et al., AJNR Am J Neuroradio/1999;20:1233-8). Direct effects of inhalation of 5% C0₂ as a vasodilativestimulus have revealed global CBF increases of approximately 87%(Kastrup A eta/., Magn Reson Imaging 2001; 19:13-20). The 60% CBFchanges shown here at two hours post ingestion of the high flavanoldrink are of a similar order to these effects.

Example 2: Evaluation of the Behavioral Effects of Cocoa PolyphenolsMethods Study Design

The study followed a double-blind, placebo-controlled, balancedcrossover design and examined the cognitive effects of multi-dose ofpolyphenols in a cocoa drink. Participants (N=30; 13 male and 17 female,mean age 21.93 years, SEM 0.61, range 18-35 years) were healthy youngadults who were screened for appropriate, potentially compromisingrelevant health conditions and dietary factors. The participants in thestudy visited the laboratory on five occasions. The first visitcomprised of a practice day where subjects were familiarized with theprocedure, this visit also served to ensure that all performance scoreswere within norms for the batteries. Subsequent visits made up the fourstudy days that were conducted not less than 3 days apart to ensure asufficient wash out between conditions, where the subjects were randomlyallocated to a Latin Square cell dictating treatment order.

Participants abstained from caffeine and alcohol for a minimum of 12hours prior to the first testing session and throughout the morninguntil the final testing session was completed. A diary was provided toallow participants to record all food and drink consumption for 24 hoursprior to the first test session of each study day. It was recommended toparticipants that they avoid food and beverages which are high inflavonoid content for 24 hours preceding each study day.

Treatments

Participants received three drinks containing: 1) 36.4 mg cocoapolyphenols (control), 2) 469.3 mg cocoa polyphenols (medium CP), 3)902.2 mg polyphenols (high CP) on separate occasions. In each case thetreatment was made up of two sachets of powder mixed with 200 ml hotwater. As well as 33% cocoa powder the drinks also contained thefollowing: non-fat dry milk powder (59.0%); fiber (5.6%); emulsifier(0.8%); cellulose gel (0.55%); xanthan gum (0.55%); artificial andnatural vanilla (0.1%); and sucralose (0.1%). Five minutes was allowedfor drink consumption.

The composition of each treatment or control sachet was as follows:

TABLE 3 CP composition HIGH CP Control % % Monomers 24.8 27 Epicatechin18.6 10.1 catechin 6.2 16.9 dimers 18.3 22.2 trimers 14.2 7.9 tetramers12.7 5.3 pentamers 9.8 3.9 hexamers 8.3 5.7 heptamers 3.8 4.7 odamers 35 nonamers 4.3 11.9 decamers 0.8 6.4 total 100 100

TABLE 4 High CP product Control product Packet code 852 217 Packet size,g 31 31 Mg CP 451.1 18.2 Calories 118.7 117.2 Total fat, g 1.4 1.5 Sat.fat, g 0.8 0.8 Cholesterol, mg 4.4 4.9 Sodium, mg 105.1 155.0 TotalCarbo., g 17.1 16.5 Dietary Fiber, g 3.0 3.9 Sugars, g 9.4 9.2 Protein,g 9.4 9.4 mg caffeine 18.3 21.2 mg theobromine 336.5 327.4

TABLE 55 Minerals High CP product Control product mg/lg mg/g Sodium, mg105.1 155.0 Potassium, mg 530.1 644.8 Calcium, mg 243.7 241.2 Iron, mg1.9 2.9 Phosphorous, mg 280.2 265.4 Magnesium, mg 85.9 78.4 Zinc, mg 1.61.6 Copper, mg 0.4 0.4 Manganese, mg 0.6 0.6

To control micro/macro-nutrients and alkaloids, two packets were fedeach time; for the high CP drink, two packets of high CP product weregiven; for the medium CP, one packet of high CP product and one packetof the control product were given; and for the control CP, two packetsof the control product were given.

Assessment Cognitive Demand Battery

In this study we undertook our ‘Cognitive Demand Battery’, (CDB).Participants received three drinks containing (i) 36.4 mg cocoapolyphenols (control), (ii) 469.3 mg cocoa polyphenols (medium CP), and(iii) 902.2 mg cocoa polyphenols (high CP). Study days involved the CDBconsisting of Serial Threes (2 min), Serial Sevens (2 min), the RapidVisual Information Processing task (RVIP, 5 min) and a mental demandvisual analogue scale (1 min). On presenting to the laboratory a salivasample was taken for analysis of caffeine levels. On each study dayparticipants went through each of the tasks once to alleviate practiceeffects.

This was followed by a baseline session, immediately followed by theday's treatment for which up to 5 min was allowed. The end of drinkconsumption represented T=O. A period of 90 min was allowed forabsorption, following which the participants underwent the CDB six timesin immediate succession, followed by a second salivary sample fordetermination of caffeine levels. The objective of these tasks (in theCDB) was to assess the impact of treatment on continuous cognitivedemand. The overall cognitive load in the session increases asparticipants complete three such tests repeatedly for a period ofapproximately one hour.

Serial Subtractions

These tasks assess the interaction between a given intervention and‘mental demand’.

A modified computerized version of the Serial Sevens test was utilized(Haskell, C. F. et al., Psychopharmocology, 2005, 179:813-825). Theoriginal verbal Serial Sevens test has appeared in a number of forms,including as part of the Mini-Mental State Examination for dementia. Ithas been used to assess cognitive impairment during hypoglycaemia, andhas also been used to investigate the relationship between increasedblood glucose levels and cognitive performance (Kennedy D. O. andScholey, A. B., Psychopharmacology, 2000, 149:63-71; Scholey, A, B, etal., Physiology & Behavior, 2001, 73:585-592).

In the current studies computerized versions of Serial Subtractions wereimplemented, using tests of 2 minutes duration. For the Serial Sevenstask a standard instruction screen informed the participant to countbackwards in sevens from the given number, as quickly and accurately aspossible, using the numeric keypad to enter each response. Participantswere also instructed verbally that if they made a mistake they shouldcarry on subtracting from the new incorrect number. A random startingnumber between 800 and 999 was presented on the computer screen, whichwas cleared by the entry of the first response. Each three-digitresponse was entered via the numeric keypad with each digit beingrepresented on screen by an asterisk. Pressing the enter key signaledthe end of each response and cleared the three asterisks from thescreen. The task was scored for total number of subtractions and numberof errors. In the case of incorrect responses, subsequent responses arescored as positive if they are correct in relation to the new number.

The Serial Threes task is identical to Serial Sevens, except that itinvolves serial subtraction of threes.

Rapid Visual Information Processing Task

This task has been widely used to study the cognitive effects ofpsychotropic interventions. The participants monitored a continuousseries of digits for targets of three consecutive odd or threeconsecutive even digits. The digits were presented at the rate of 100per minute and the participant responded to the detection of a targetstring by pressing a response key as quickly as possible. The task wascontinuous and lasted for 5 minutes, with 8 correct target strings beingpresented in each minute. The task was scored for percentage of targetstrings correctly detected, average reaction time for correctdetections, and number of false alarms.

Subjective Scales {Mental Demand Visual Analogue Scale)

At the end of each set of tasks participants were asked to indicate howmentally fatigued they felt by marking a 100 mm line with the end-pointslabeled “not at all” and extremely.

Results

The results are shown in FIG. 4(a-d). Results show beneficial effects ofa cocoa drink supplemented with flavanols/procyanidins on certainaspects of cognitive performance/abilities.

Example 3: Assessment of Acute Behavioral Effects of Cocoa Polyphenolson Mentally Non-Demanding Tasks Method

This study allowed exploration of the effects of cocoa polyphenolsindependent of mentally-demanding situations and allowed to assesswhether the effects of cocoa polyphenols were restricted solely tomentally-demanding situations.

Prior to participation in the study volunteers signed an informedconsent form and completed a medical health questionnaire. Allparticipants reported that they were in good health and free from socialdrugs and medication, with the exception of the contraceptive pill.Habitual smokers were excluded from the study.

Thirty participants were recruited into the study and informed that itsaim was to investigate the cognitive and mood effects of a cocoa drinkcontaining active components (one of which may be caffeine).Participants abstained from caffeine and alcohol for a minimum of 12hours prior to the first testing session and throughout the morninguntil the final testing session was completed. A diary was provided toallow participants to record all food and drink consumption for 24 hoursprior to the first test session of each study day and throughout thestudy visit. It was recommended to participants that they avoid food andbeverages which are high in flavonoid content for 24 hours precedingeach study day. This request for restricted flavonoid diet allowed acutechanges in plasma flavanollevels to be monitored.

Salivary Caffeine Levels

Saliva samples were obtained using salivettes. Samples were takenimmediately following baseline assessment in order to confirm complianceto overnight abstinence and immediately following each post-treatmentassessment session to confirm uniform caffeine absorption acrossconditions. The saliva samples were immediately frozen at −20° C. untilthawing for in-house batch analysis using the Emit system (Syva, PaloAlto, USA). This is an enzyme immunoassay intended to measure caffeineas a metabolite and is based on competition for antibody binding sitesbetween caffeine and an enzyme labelled drug.

Plasmajlavanollevels

Immediately prior to the baseline assessment and prior to the 90 minutepost-treatment session a 2 ml venous blood sample was taken fordetermination of flavanollevels. These blood samples were collected byvenipuncture using a monovette containing EDTA. The samples were thenkept on ice until centrifuged at 3,000 rpm for 10 minutes at atemperature of 5° C. A Gilson pipette was used to measure 1485 Jll ofthe resulting plasma into an amber tube. A fresh solution of ascorbicacid (570 mM/100.4 mg/ml) was also prepared, kept on ice and away fromlight. 15 Jll of this solution was then added to the 1485 J.ll ofplasma, mixed and stored at −70° C. Unfortunately due to technicaldifficulties analyses of these samples was not possible.

Assessment Cognitive Drug Research (CDR) Battery

The CDR system has been used in well over 500 European and NorthAmerican drug trials, and has been shown to be sensitive to cognitiveimprovements as well as impairments with a wide variety of substances.

A tailored version of the battery was used, similar to that which haspreviously been found to be sensitive to improved cognitive function asa consequence of ingestion of numerous nutraceuticals. The selection ofcomputer controlled tasks from the system was administered with parallelforms of the tests being presented at each testing session. Presentationwas via color monitors on laptops, and, with the exception of writtenword recall tests, all responses were recorded via two-button (YES/NO)response boxes.

The entire selection of tests took approximately 20 minutes and were.administered in the following order:

Word Presentation: Fifteen words, matched for frequency andconcreteness, were presented in sequence on the monitor for theparticipant to remember. Stimulus duration was 1 second, as was theinter-stimulus interval.

Immediate Word Recall: The participant was allowed 60 seconds to writedown as many of the words as possible. The task was scored for numbercorrect and errors.

Picture Presentation: Twenty photographic images for the participant toremember were presented sequentially on the monitor, at the rate of 1every 3 seconds, with a stimulus duration of 1 second.

Simple Reaction Time: The participant was instructed to press the ‘YES’response button as quickly as possible every time the word ‘YES’ waspresented on the monitor. Fifty stimuli were presented with aninter-stimulus interval that varies randomly between 1 and 3.5 seconds.Reaction times were recorded in milliseconds.

Digit Vigilance Task: A target digit was randomly selected andconstantly displayed to the right of the monitor screen. A series ofdigits was presented in the center of the screen at the rate of 80 perminute and the participant was required to press the ‘YES’ button asquickly as possible every time the digit in the series matches thetarget digit. The task lasted three minutes and there was 45stimulus-target matches. Task measures were accuracy (%), reaction time(msec) and number of false alarms.

Choice Reaction Time: Either the word ‘NO’ or the word ‘YES’ waspresented on the monitor and the participant was required to press thecorresponding button as quickly as possible. There were 50 trials, ofwhich the stimulus word was chosen randomly with equal probability, witha randomly varying inter-stimulus interval of between 1 and 3.5 seconds.Reaction times (msec) and accuracy (%) were recorded.

Spatial Working Memory: A pictorial representation of a house waspresented on the screen with four of its nine windows lit. Theparticipant was instructed to memorize the position of the illuminatedwindows. In 36 subsequent presentations of the house, one of the windowswas illuminated and the participant decided whether or not this matchedone of the lighted windows in the original presentation. The participantmade their response by pressing the ‘YES’ or ‘NO’ response button asquickly as possible. Mean reaction times were measured in msec, andaccuracy of responses to both original and novel (distractor) stimuliwere recorded as percentages.

Numeric Working Memory: Five digits were presented sequentially for theparticipant to hold in memory. This was followed by a series of 30 probedigits for each of which the participant decided whether or not it hadbeen in the original series and pressed the ‘YES’ or ‘NO’ responsebutton as appropriate as quickly as possible. This was repeated twofurther times with different stimuli and probe digits. Mean reactiontimes were measured in msec, and accuracy of responses to both originaland novel (distractor) stimuli were recorded as percentages.

Delayed Word Recall: The participant was again given 60 seconds to writedown as many of the words as possible. The task was scored as numbercorrect and errors.

Delayed Word Recognition: The original words plus 15 distractor wordswere presented one at a time in a randomized order. For each word theparticipant indicated whether or not she recognized it as being includedin the original list of words by pressing the ‘YES’ or ‘NO’ button asappropriate and as quickly as possible. Mean reaction times weremeasured in msec, and accuracy of responses to both original and novel(distractor) stimuli were recorded as percentages.

Delayed Picture Recognition: The original pictures plus 20 distractorpictures were presented one at a time in a randomized order. For eachpicture participants indicated whether or not it was recognized as beingfrom the original series by pressing the ‘YES’ or ‘NO’ button asappropriate and as quickly as possible. Mean reaction times weremeasured in msec, and accuracy of responses to both original and novel(distractor) stimuli were recorded as percentages.

The following tables indicate which stage of information processing eachtask assesses:

TABLE 6 Level 1: Attention The ability to select, evaluate and respondto appropriate environmental information Simple Reaction Time AlertnessPower of concentration Primary stage of information processing ChoiceReaction Time As above, plus Stimulus discrimination Responseorganization Task Cognitive States and Processes Assessed DigitVigilance Intensive vigilance Sustained concentration Ability to ignoredistraction

TABLE 7 Level II: Short Term or Working Memory The ability totemporarily store the information relevant to ongoing tasks TaskCog11itive States and Processes Assessed Digit Scanning Sub-vocalrehearsal of digit sequences Articulatory loop sub-system of workingmemory Spatial Working Ability to temporarily retain spatial informationMemory Visuo-Spatial sub-loop of working memory

TABLE 8 Level III: Long Term or Episodic Secondary Memory The ability toregister, store and retrieve information over any period required TaskCognitive States and Processes Assessed Word Recall Ability to store andrecall verbal information Capacity for un-cued retrieval of wordsEpisodic secondary verbal recall Word Recognition Ability (speed andsensitivity) to discriminate novel from previously presented wordsEpisodic secondary verbal recognition Picture Recognition Ability todiscriminate novel from previously presented pictorial informationEpisodic secondary non-verbal visual recognition

Bond-Lader Visual Analogue Scales (Bond A., and Lader M., 1974, The useof analogue scales in rating subjective feelings, British Journal ofPsychology, 47:211-218).

This measure was made up of 16 visual analogue scales with the endpoints anchored by antonyms: alert-drowsy, calm-excited, strong-feeble,muzzy-clearheaded, well coordinated-clumsy, lethargic-energetic,contented-discontented, troubled-tranquil, mentally slow-quick witted,tense relaxed, attentive-dreamy, incompetent-proficient, happy-sad,antagonistic-friendly, interested-bored, withdrawn-sociable. These werecombined to form three “mood” factors: ‘alert’, ‘calm’ and ‘content’.

Psychomotor Vigilance Task (Dinges D. F., and Powell J. W., 1985,Microcomputer analyses of performance on a portable, simple visual RTtask during sustained operations. Behavior Research, Methods,Instruments and Computers, 17:652:655).

The psychomotor vigilance task (PVT) is a simple portable reaction timetask used to evaluate sustained attention. The subject was instructed topress a button with the thumb of their dominant hand as soon as thestimulus appears (LED-digital counter). In the present study, theduration of a single PVT trial comprised 10 min. Sleepiness was alsorated with this device before and after the reaction time task with theuse of a 10-point scale.

Treatments

Participants received three drinks containing: 1) 36.4 mg cocoapolyphenols (control), 2) 469.3 mg cocoa polyphenols (medium CP), 3)902.2 mg polyphenols (high CP) on separate occasions. In each case thetreatment was made up of two sachets of powder mixed with 200 ml hotwater. The control treatment consisted of 2 sachets of control CP, the469.3 mg dose consisted of one control sachet plus one high CP sachet,and the 902.2 mg dose consisted of 2 high CP sachets. Nutritionalinformation for the 2 sachets can be found in Tables 3-5 of Example 2.Five minutes was allowed for drink consumption.

Each participant was required to attend a total of four study days thatwere conducted not less than 3 days apart to ensure a sufficient washout between conditions. Testing took place in a suite of laboratorieswith participants visually isolated from each other. On arrival at theirfirst session on the first day participants were randomly allocated to atreatment regime using a Latin square design which counterbalanced theorder of treatments across the three active days of the study. The firstday involved completion of the test battery four times. This wasundertaken in order to control for practice effects and to allowfamiliarization with the test battery and procedure on subsequentvisits. The practice day data were not included in any analyses.

Each of the three active study days comprised four identical testingsessions. The first was a pre-dose testing session, which establishedbaseline performance for that day. This was followed immediately byingestion of that day's treatment. Further testing sessions began at 90minutes, 3 hours and 6 hours following consumption of the day'streatment. Each testing session lasted approximately 30 minutes andcomprised completion of the CDR test battery, Bond-Lader visual analoguemood scales, 10-minute PVT and production of a saliva sample with theuse of a salivette. In addition, the pre-dose session and the 90 minutepost-dose session also involved the taking of a 2 ml venous plasmasample prior to completion of the CDR battery.

Salivary caffeine levels were analysed to assess compliance to caffeineabstinence.

Prior to the primary statistical analysis separate, one way, repeatedmeasures ANOVAS of pre-dose baseline data were conducted to ascertainany chance baseline differences in performance prior to the treatments.

Scores on the individual task outcomes were analysed as ‘change frombaseline’ using SPSS 12.0.1.

The data from each measure were analysed by two-way repeated measuresANOVA [time (1.5, 3 and 6 hours post-dose) X treatment (469.3 mgCP/902.2 mg CP/control)].

Results

Mean pre-dose baseline, and change from baseline scores for each measurefor each condition were determined along with F-values and probabilitiesfor effects of treatment.

Prior to analysis of change from baseline data, mean pre-dose rawbaseline scores for all three conditions (control, 469.3 mg CP, 902.2 mgCP) for each outcome measure were subjected to a one-way,repeated-measures ANOVA. There were no significant baseline differencesfor any measure.

Salivary analysis confirmed adherence to caffeine abstinenceinstructions with mean baseline caffeine values of 0.79 μg/ml (levelsjust below 1 μg/ml have been reported for overnight caffeineabstinence-Evans and Griffith, 1999, Caffeine withdrawal: a parametricanalysis of caffeine dosing conditions. The Journal of Pharmacology andExperimental Therapeutics 289:285-294). Analysis of post-treatmentsalivary caffeine levels revealed no significant differences betweentreatment conditions.

Post-Treatment Scores

There were no significant post-treatment differences for any measure.

The tasks which make up the CDR assessment battery are wide-ranging inthe abilities they measure but they are simple tasks which do notrequire a lot of effort. The results from this study show no effect ofthe compounds of the invention during performance of mentallynon-demanding tasks.

What is claimed is:
 1. A method of enhancing executive cognitivefunction in a subject in need thereof comprising administering to thesubject a composition comprising an effective amount of at least onecompound selected from the group of catechin, epicatechin, a compoundhaving the formula A_(n), a pharmaceutically acceptable salt thereof,and a derivative thereof:

wherein n is an integer from 2 to 18; R and X each have either α or βstereochemistry; R is OH, O-sugar or O-gallate; the substituents of C-4,C-6 and C-8 are X, Z and Y, respectively, and bonding of monomeric unitsoccurs at C-4, C-6 or C-8; when any C-4, C-6 or C-8 is not bonded toanother monomeric unit, X, Y and Z are hydrogen or a sugar; and thesugar is optionally substituted with a phenolic moiety at any position,for instance, via an ester bond.
 2. The method of claim 1, wherein R isOH.
 3. The method of claim 2, wherein the subject is a human.
 4. Themethod of claim 2, wherein the subject is a veterinary animal.
 5. Themethod of claim 3, wherein the compound is catechin.
 6. The method ofclaim 3, wherein the compound is epicatechin.
 7. The method of claim 3,wherein the compound is A_(n), and n is 2 to
 12. 8. The method of claim3, wherein the compound is A_(n) and n is 2 to
 10. 9. The method ofclaim 3, wherein n is
 2. 10. The method of claim 3, wherein thecomposition is a pharmaceutical composition.
 11. The method of claim 3,wherein the composition is a food.
 12. The method of claim 3, whereinthe composition is a beverage.
 13. The method of claim 3, wherein thecomposition is a confectionery.
 14. The method of claim 11, wherein thecompound is provided as a cocoa extract.
 15. The method of claim 11,wherein the compound is provided as a cocoa ingredient.
 16. The methodof claim 3, wherein the human is selected from the group of: a humanparticipating in a competitive event, a human addressing an audience orgathering, and a human in entertainment profession.
 17. The method ofclaim 3, wherein the human is selected from the group of: a humanperforming complex decision-making and/or multitasking, a transportationworker, a construction worker, and defense personnel.
 18. The method ofclaim 3, wherein the human is an elderly individual suffering from adecline in cognitive abilities.
 19. The method of claim 3, wherein thehuman is healthy.
 20. A method of increasing blood flow in brainvasculature in a healthy subject in need thereof comprisingadministering to the subject a composition comprising an effectiveamount of at least one compound selected from the group of catechin,epicatechin, a compound having the formula A_(n), a pharmaceuticallyacceptable salt thereof, and a derivative thereof:

wherein n is an integer from 2 to 18; R and X each have either α or βstereochemistry; R is OH, O-sugar or O-gallate; the substituents of C-4,C-6 and C-8 are X, Z and Y, respectively, and bonding of monomeric unitsoccurs at C-4, C-6 or C-8; when any C-4, C-6 or C-8 is not bonded toanother monomeric unit, X, Y and Z are hydrogen or a sugar; and thesugar is optionally substituted with a phenolic moiety at any position,for instance, via an ester bond.